Conjugated linoleic acid (CLA) is a mixture of positional and geometric isomers of linoleic acid which is found preferentially in dairy products and meat. Preliminary studies indicate that CLA is a powerful anticarcinogen in the rat mammary tumor model with an effective range of 0.1 - 1% in the diet. This protective effect of CLA is noted even when exposure is limited to the time from weaning to carcinogen administration. The timing of this treatment corresponds to maturation of the mammary gland to the adult stage, suggesting that CLA may have a direct effect in reducing cancer risk of the target organ. The hypothesis being tested is that the selective incorporation of a particular isomer of CLA (c9,t11) into membrane phospholipids may change the responsiveness of mammary epithelial cells (MEC) to growth modulatory factors. This in turn may affect the proliferation, morphogenesis, differentiation and transformation of the target cells. Additionally, CLA is a potent antioxidant. Since adipocytes are an integral component of the mammary gland, accumulation of CLA (all isomers) in triglycerides of the fat cells may bestow some insulation against oxidant stress in the microenvironment of the epithelial cell. Both animal bioassay and cell culture will used for the proposed research. AIM 1. To delineate the role of CLA in initiation versus post-initiation stages of mammary carcinogenesis induced by either methylnitrosourea (MNU) or 7,12-dimethylbenz(a)anthracene (DMBA). These studies are designed to further characterize the cancer protective action of CLA and to determine whether continuous feeding of CLA is necessary to produce a maximal inhibitory response. AIM 2. To investigate whether the fat content of a diet (both level and type) will modulate the efficacy of CLA in cancer prevention. This issue is critical in providing insight regarding the impact of CLA as an anticarcinogen in populations with a high fat intake. AIM 3. To examine the incorporation of CLA into membrane phospholipids of MEC and into triglycerides of mammary fat cells. The goal is to investigate the regulation of CLA incorporation by environmental and host factors and the significance of this accumulation in conferring resistance to mammary gland carcinogenesis. AIM 4. To delineate the effects of CLA on proliferation as well as morphological and functional differentiation of normal rat MEC (RMEC) in primary culture and to determine whether CLA alters their hormonal sensitivity. The effect of CLA will be evaluated by direct addition to the culture or by using RMEC isolated from CLA-fed rats. AIM 5. To determine whether CLA alters the sensitivity of RMEC to in vitro transformation and/or whether it alters the growth of transformed RMEC. AIM 6. To determine whether CLA alters stromal-epithelial interactions in the mammary gland and the functional consequence of such an effect. To this end, mammary stromal cells (adipocytes, fibroblasts) isolated from CLA-fed rats, or cultured with CLA in vitro, will be co-cultured with normal or carcinogen-treated RMEC and a determination made as to whether CLA treatment modulates the efficacy of the stromal cells in modifying the proliferation, differentiation or transformation of RMEC in primary culture